KIR3DS1 is presumed to be an NK cell activation receptor. The gene has many (>40) inhibitory alleles, known as KIR3DL1. Various KIR3DL1 subtypes have been shown to interact directly with HLA-I proteins with the Bw4 public epitope. Regardless of its similarity to KIR3DL1, and its established role in HIV disease, KIR3DS1 has only recently been shown to be expressed on NK cells and no ligand has been described. Toward an understanding of KIR3DS1 ligand binding we have been conducting studies using Bw4 HLA-I tetramers folded with various peptides. These studies have, thus far, demonstrated no binding to KIR3DS1. Therefore, we have used several new alleles of KIR3DL1 as a guide for a mutagenic study of KIR3DS1. In this work specific residues of KIR3DS1 are mutated to match various residues of KIR3DL1. We evaluate the ability of KIR3D specific antibodies and HLA tetramers to recognize these new KIR3DL1 species and superimpose these findings on the presumed three dimensional structure of KIR3DL1. Through this analysis we can define specific sites on KIR3DS1 that control the ligand binding specificity and draw conclusions regarding its ligand. The high frequency of KIR3DS1 we have previously described, together with this receptors ability to activate NK cells and be maintained during HIV viremia, suggest that our dissection of its binding characteristics will lead to exciting new approaches to HIV therapy.